Costimulatory genes and molecules have a profound impact on autoimmune diseases and recent data suggests that these molecules may control genetic susceptibility to many autoimmune diseases. Of the costimulatory receptors, CTLA4 has gained considerable attention because loss of CTLA4 results in multi-organ autoimmune diseases in mouse. Linkage to CTLA4 in mouse and man has been found due to generation of alternatively spliced forms of CTLA4. In mouse, the type 1 diabetes is linked to the production of a novel isoform of CTLA4 named ligand independent CTLA4 (HCTLA4), and in human autoimmune diseases the susceptibility is linked to the soluble form of CTLA4 (sCTLA4). We have recently generated a transgenic mouse that expresses HCTLA4 in all T cells and when crossed onto the CTLA4-/- mice, the HCTLA4 transgenic expression was able to rescue the hyperproliferative phenotype and early lethality in the CTLA4 -/- mice. These data raise the issue of how UCTLA4 maintains self-tolerance and what is the mechanism by which sCTLA4 regulates autoimmunity in humans. To address these issues we have proposed the following aims: 1. Study the role of UCTLA4 splice variant in the development of peripheral tolerance. We will determine why HCTLA4 expression prevents the autoimmune disease in the CTLA4-/- mice, whether HCTLA4 inhibits activation with the low affinity but not high affinity T cell ligands and what its role might be in the induction of high dose versus mucosal tolerance; 2. Study the role of sCTLA4 in the regulation of autoimmune responses and development of autoimmune disease. We will generate transgenic mice, which will express sCTLA4 in T cells. These transgenic mice will be utilized for the testing the function of sCTLA4 in the development of diabetogenic repertoire and in the development of diabetes and EAE in mice; 3. Study the role of PTA-1. a Th1 specific costimulatory molecule, in the regulation of autoreactive T cell repertoire and autoimmune disease. We have recently discovered that PTA-1 (CD226) is specifically expressed on Th1 and its expression shows loss on Th2 cells upon differentiation. In vivo administration of an antibody to PTA-1 results in the inhibition of T cell response and affecting both T cells and APC function. Using the antibody to this Th1 specific molecule we will determine the relative role of PTA-1 on T cells and its role on APCs in costimualting T cell activation. These studies will provide fundamental information on the mechanism by which the negative costimualtory molecules associated with genetic susceptibility play a role in the regulation of autoimmune diseases.